Immobilization Of Heparin To Collagen Matrix And Its Application For Transforming Growth Factor (TGF) Controlled Release

The chemical modification on collagen matrix by crosslinking and heparinization is toimprove mechanical properties, controlled degradation rate and the release of growth factor.The modified collagen matrices mimic the extracellular matrices in the structure,compositionand biological behavior. The crosslinked and heparinized collagen matrices, load bone growthfactor (TGF), are expected to be the scaffolds for cartilage tissue engineering in cartilagerepair.Collagen extracted from the ox tendon is made into gel, collagen gel is mixed withamountsof chitosan. After freeze drying at-80℃, the collagen/chitosan sponge was obtained.Collagen/chitosan sponge was treated with1-(3-Dimethylaminopropyl)-3-ethyl-carbodiimide/N-Hydroxysuccinimide, Genipin andheparin. The modified collagen matrices were characterized by scanning electron microscope,crosslinking degree, water absorption, mechanical strength, in vitro degradation, cytotoxicityand proliferation. We compared the differences physicochemical properties of each group,including the untreated control group, EDC/NHS group, GP group and EDC/NHS-Hep group,GP-Hep group.Toluidine blue was used to measure heparin content in collagen/chitosan matrices, thecontent of heparin increases with the content of chitosan in the collagen matrix. Through the2,4,6-Trinitrobenzenesulfonic acid solution method in the measurement of the free aminogroups, it is found that free amino groups of EDC/NHS group, GP group, EDC/NHS-Hepgroup, GP-Hep group decreased significantly compared with the control group. In collagengroups, the water absorption capacity of EDC/NHS group, GP group, EDC/NHS-Hep group,GP-Hep group was greatly improved after crosslinking, while in Col/Cht groups the waterabsorption capacity of EDC/NHS group, EDC/NHS-Hep group was decreased. Those of GPgroup and GP-Hep group were slightly improved. The water absorption of EDC/NHS groupand GP group were higher than that of EDC/NHS-Hep group or GP-Hep group. The results ofdegradation experiment in vitro showed that the stability against enzymatic digestion wasimproved. After25days degradation in vitro, the degradation rate of EDC/NHS group andEDC/NHS-Hep group was between30%~40%, but the degradation rate of GP group andGP-Hep group was lower than20%, while the control group was almost completely degradedwithin15days. The cytotoxicity was detected by MTT and the results implied that the celltoxicity of each group was scarely observed.Using double emulsion method, the PLGAmicrospheres loading TGF were prepared with the TGF entrapment efficiency of about70%.The size of most particles is less than2μm, among which about60%was less than1μm.Through45days of release experiment in vitro, it is found that the PLGA microspheres couldslower the release of the polypeptide TGF in a long time. The release of TGF is stable usingcollagen matrix loaded PLGA microspheres. It can effectively prolong the time of drug release, as compared with, the TGF-PLGA microspheres.